The present invention relates to a method for screening drugs for use in treating hypertension using the tubular renin-angiotensinogen system identified by the present invention. The invention further relates to a method to diagnose sodium status in an individual by measuring urinary angiotensinogen, angiotensin-I, des-AI-angiotensinogen or renin.
The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference, and for convenience are referenced in the following text by author and date and are listed alphabetically by author in the appended list of references.
The following abbreviations are used herein: A-I-angiotensin-I; A-II-angiotensin-II; ACE-angiotensin converting enzyme; AGT-angiotensinogen gene; ANG or Ang-angiotensinogen protein; -6(A)/-6(G)-promoter polymorphism at position -6; CCD-cortical collecting duct; CNT cortical connecting tubule; DCT-distal convoluted tubule; IC-intercalated cells; JGA-juxtaglomerular apparatus; PCR-polymerase chain reaction; RAS-renin-angiotensin system; RT-PCR-reverse transcriptase polymerase chain reaction; and HPLC-high pressure liquid chromatography.
Blood pressure control is intrinsically linked to fluid volume balance and electrolyte homeostasis. Regulation of plasma volume in response to variation in dietary sodium (1) is primarily controlled by the renin-angiotensin system (RAS) and its main effector angiotensin-II (A-II); this peptide hormone is released from angiotensinogen (Ang) by two cleavage steps involving renin and angiotensin-converting enzyme (ACE) (2).
The short-term effects of A-II are better understood than its long-term effects. Acute depletion of body fluid volume triggers a vasoconstrictor response mediated by the circulating renin-angiotensin system (RAS), involving renin secreted by the juxtaglomerular apparatus (JGA) in the kidney, Ang from liver, and ACE present in the luminal cell membrane of capillary endothelium.
Sustained low-dose infusion of A-II leads to progressive, long-term elevation of arterial pressure due to cumulative sodium retention primarily mediated by direct intrarenal A-II effects1. A-II has been detected in proximal tubular luminal fluid at high concentrations (3, 4). In contrast to plasma renin (36-40 kDa), Ang (61-65 kDa) is not filtered through the glomerular basement membrane. Detection of abundant angiotensinogen mRNA in proximal tubule epithelium (5-7), strongly suggests local generation of A-II at this site by an as yet unspecified mechanism. Renin mRNA can be detected in proximal tubule only by application of the very sensitive technique of RT-PCR (8). Exogenous A-II stimulates the luminal sodium-hydrogen exchanger present in the proximal tubule cells (9, 10) and also stimulates epithelial sodium channels and possibly other transporters in the distal segments of nephron (11-14).
Fundamental questions remain unanswered, however. If intrarenal A-II directly affects sodium reabsorption, where is it generated, and by what mechanism? How is this mechanism regulated in response to sodium? At what sites does A-II impact on sodium transport along the nephron? What is the mechanism for coordinated regulation of sodium uptake in proximal and distal segments of the nephron? Can it allow for a decoupling of sodium reabsorption and potassium excretion in the distal tubule?
It is desired to address these questions and to elucidate answers which can be used for screening drugs and diagnosing sodium status of an individual.